Chronic lymphocytic leukemia (CLL) is the most common form of leukemia in the United States. However, an understanding of the molecular abnormalities which underlie this disease is lacking. CLL is characterized by the continuous unrestrained growth of B lymphocytes. As such, a defect in a signaling pathway that controls the regulation of mitosis in B lymphocytes might be responsible for this malignancy. The proliferation of B lymphocytes is normally closely regulated by a family of secreted cytokines which act by binding to specific cell surface receptors. Upon binding to their cognate receptor, most cytokines signal, at least in part, by activating a set of transcription factors known as STATs. STATs can be phosphorylated on a single tyrosine residue, which leads to STAT dimerization, nuclear translocation, DNA binding, and transcriptional activation. In addition, the function of several STATs is modulated by phosphorylation on specific serine residues. This additional serine phosphorylation appears to be important for gene activation, and may serve to integrate signals generated by diverse intracellular pathways. Given the central role of STATs in modulating the growth and function of B lymphocytes, we considered the possibility that abnormalities in STAT-mediated signaling might underlie the defective growth regulation seen in the malignant B lymphocytes of CLL. We have recently reported that although tyrosine phosphorylation of STATs is not present in CLL, constitutive phosphorylation of STAT1 and STAT3 on specific serine residues, ser-727 in each protein, is found in the peripheral blood B lymphocytes of all patients with CLL examined thus far, but not in normal B cells. The present proposal seeks to extend these findings by exploring the mechanism of this phosphorylation and its consequence in the biology of CLL. Four specific aims will be pursued: (1) To analyze the effect that serine phosphorylation of STAT1 and STAT3 plays in mediating gene activation and cell survival in CLL cells and normal lymphocytes; (2) To determine the effect that inhibition of STAT tyrosine phosphorylation has on the biology of transformed B lymphocytes; (3) To analyze the effect that a chemotherapeutic drug used to treat CLL, fludarabine, has on STAT signaling; and (4) To isolate the kinase(s) responsible for the phosphorylation of ser-727 of STAT1 and STAT3 in CLL cells. As a result of these experiments, we plan to discern the role of STATs in the abnormal growth of CLL cells, and to identify targets for novel and specific therapies.